This invention relates to a method for determining the quantitative levels of serum or tissue protein. More particularly, this invention concerns itself with the quantitative determination of protein levels by measuring the changes in fluorescence emission of an immunospecific affinity fluorescent agent which is specifically induced by the molecular interactions between a target protein and the fluorescent agent.
At the present time, a significant need exists for a versatile, accurate, rapid, and economical test to quantitate levels of serum and tissue proteins, particularly human immunoglobulin. Radial immunodiffusion, solid-phase immunosorbent, and radioimmuno assays are conventional techniques presently utilized. Although each of these have proven somewhat successful, they all suffer certain disadvantages. In attempting to improve on these prior art methods, various alternative techniques are continually being developed and refined. For example, immunoenzymatic amplification and laser doppler electrokinetics are recent methods conceived for improving protein determinations.
As is well known, fluorescent probes constitute a very useful tool in the study of protein structures because changes in the conformation of the protein conjugate may significantly alter the emission properties of the probe. As a consequence, a research effort has evolved which is directed towards the goal of developing a method for determining protein level concentrations through the use of fluorescent type indicators. The resulting research effort produced the novel immunospecifically-directed fluorescent enhancement technique of this invention which utilizes N-(3-pyrene) maleimide, hereinafter referred to as PM, as the immunospecific fluorescent reagent or fluorescent probe. This novel method provides a simple, rapid and inexpensive, quantitative technique for determining concentration levels of serum or tissue protein, particularly human immunoglobulin, hereinafter referred to as IgG.